Sperm traits on in vitro production (IVP) of bovine embryos: Too much of anything is good for nothing

Siqueira, Adriano Felipe Perez; Castro, Luiz Guilherme Martins; Assis, Patrícia Monken de; Bicudo, Luana de Cássia; Mendes, Clayton Quirino; Nichi, Marcílio; Visintin, José Antônio; Assumpção, Mayra Elena Ortiz D’Ávila

doi:10.1371/journal.pone.0200273

Cited by 18 publications

(19 citation statements)

References 52 publications

(64 reference statements)

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“…In other words, when a population of sperm cells appears to contain a relatively denser head, they are also more difficult to capacitate. Another interpretation of our findings is that more massive heads are harder to “push” with smaller tails, which is consistent with findings concerning head shape ( 51 , 52 ). Likewise, it was observed that deer sperm with longer midpieces swim at an overall slower speed ( 53 ).…”

Section: Discussionsupporting

confidence: 92%

Reproductive outcomes predicted by phase imaging with computational specificity of spermatozoon ultrastructure

Kandel

Rubessa

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

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The ability to evaluate sperm at the microscopic level, at high-throughput, would be useful for assisted reproductive technologies (ARTs), as it can allow specific selection of sperm cells for in vitro fertilization (IVF). The tradeoff between intrinsic imaging and external contrast agents is particularly acute in reproductive medicine. The use of fluorescence labels has enabled new cell-sorting strategies and given new insights into developmental biology. Nevertheless, using extrinsic contrast agents is often too invasive for routine clinical operation. Raising questions about cell viability, especially for single-cell selection, clinicians prefer intrinsic contrast in the form of phase-contrast, differential-interference contrast, or Hoffman modulation contrast. While such instruments are nondestructive, the resulting image suffers from a lack of specificity. In this work, we provide a template to circumvent the tradeoff between cell viability and specificity by combining high-sensitivity phase imaging with deep learning. In order to introduce specificity to label-free images, we trained a deep-convolutional neural network to perform semantic segmentation on quantitative phase maps. This approach, a form of phase imaging with computational specificity (PICS), allowed us to efficiently analyze thousands of sperm cells and identify correlations between dry-mass content and artificial-reproduction outcomes. Specifically, we found that the dry-mass content ratios between the head, midpiece, and tail of the cells can predict the percentages of success for zygote cleavage and embryo blastocyst formation.

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Section: Discussionsupporting

confidence: 92%

Reproductive outcomes predicted by phase imaging with computational specificity of spermatozoon ultrastructure

Kandel

Rubessa

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

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“…It is possible to transfer the whole oocyte to the laboratory or do aspiration of visualized follicles and transfer them in follicular fluid with oocyte-cumulus complexes in buffered medium. Follicles of from 2 to 8 mm are aspirated with a 18G needle attached to a 5-10 ml syringe [18,24]. Transportation is done in special incubators with constant temperature 30-35 °С [11,24].…”

Section: Sampling and Maturation Of Oocytesmentioning

confidence: 99%

“…Follicles of from 2 to 8 mm are aspirated with a 18G needle attached to a 5-10 ml syringe [18,24]. Transportation is done in special incubators with constant temperature 30-35 °С [11,24]. According to the data obtained by L. V. Golubets and co-authors, the optimum delivery time of oocytes tissue or aspirated oocyte-cumulus complexes at temperature of 37 °С is not more than 3 hours, whereas formation of morulas/ blastocytes can be up to 27 %.…”

Section: Sampling and Maturation Of Oocytesmentioning

confidence: 99%

“…Main modifications are caused by changes in concentration of gonadotropins introduced into culture medium. As a rule, doses of 0,5 ug/ml FSH (follicle stimulating hormone) and 5-50 ug/ml HGG (human chorionic gonadotropin) or 5 ug/ml LH (luteinizing hormone) are used [4,12,22,24]. Methods of cultivation of donor oocytes in medium with prolactin are developed and successfully used [5,6] Preparation of spermatozoids.…”

Section: Sampling and Maturation Of Oocytesmentioning

confidence: 99%

“…Amount of sperm added to oocyte-cumulus complexes during in vitro fertilization affects percentage of zygotes inseminated by a few spermatozoids (polyspermy). As a rule, it is enough to introduce in the medium sperm in concentration of about 1.0-9.0×10 6 mobile spermatozoids/ml [19,24].…”

Section: Sampling and Maturation Of Oocytesmentioning

confidence: 99%

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Features of the preparation of biological material for genome editing in cattle

Баркова

Makutina²,

Modorov³

et al. 2019

Agrarian Bulletin of The

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Abstract. To carry out genome editing in cattle, an effective and well-functioning system for obtaining gametes, fertilizing eggs and their cryopreservation is necessary. Aim of the work: review and research of present-day existing methods of obtaining, insemination and cryopreservation of donor material, in order to provide genome editing in cows. Methods and materials. The work is completed according to the theme No. 0532-2019-0001 “Development of complex technology of marker-based genome selection of agricultural animals” within State Order of Ministry of Education and Science of the Russian Federation. The analysis of open scientific literature on the issues of in vitro fertilization in animals, cryopreservation of oocytes and embryons, sperm preparation and methods of insemination of cows’ oocytes, and cryopreservation of oocytes and embryons of animals is done. Features of the preparation of biological material of cattle for genome editing by microinjection into ooplasm are described. Results of research and duscussion. At present time there are two ways to obtain donor material from cattle: from live animals and taking ovaries after slaughtering cows. Material transportation is carried out at a temperature of 30–37 °C depending on the distance to the laboratory and expected time period of transportation. Oocyte-cumulus complexes can be removed by ovarian dissection and aspiration of visible follicles. In both cases, immature eggs are predominantly obtained. Subsequent ripening is carried out in vitro using special media in a CO2 incubator. The culture medium for oocyte maturation should contain hormones that mimic the peak of LH (luteinizing hormone), which occurs in vivo during the maturation of oocytes before ovulation. To accumulate a certain number of eggs at the stage of MII, it is recommended to carry out their cryopreservation by the method of vitrification, having previously released the oocyte from the cumulus cells. After thawing, oocytes need to be incubated for 2–3 hours 38.5 °C in 5–6.5% CO2 to restore the spindle. In order to make editing more effective, the introduction of genetic material is recommended to be carried out in parallel with the fertilization method “icsi”. In humans, mice and rabbits, an injection of sperm into the cytoplasm is sufficient to activate the oocyte, however, in cattle, just micro-injection of the sperm is not enough and often the male pronucleus does not form. To solve the problem, various methods are used, including freezing-thawing of sperm, resulting in damage of a membrane, or addition of heparin-glutathione into the medium that increases decondensation of the sperm DNA.

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Assessment of different sperm functional tests in golden‐headed lion tamarins (Leontopithecus chrysomelas)

Arakaki

Salgado

Losano

et al. 2019

American J Primatol

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The golden‐headed lion tamarin (Leontopithecus chrysomelas) is an endangered species endemic to Brazil's Atlantic Forest, a shrinking biodiversity hotspot. As in other Neotropical primates, its semen characteristics and freezability are poorly studied. Hence, reproductive technologies for callitrichids would greatly benefit from reliable methods of semen analysis. In a bid to promote reproductive research in tamarins, we validated simple and inexpensive sperm function tests that can be used to monitor sperm‐egg binding, plasma membrane and acrosome integrity, mitochondrial activity, and DNA fragmentation. Ejaculates from adult males were individually diluted and divided into control and damage‐induced aliquots, and then samples comprising assorted amounts of damaged spermatozoa were examined by organelle‐specific tests. Our findings showed that sperm‐binding in chicken egg perivitelline membrane (EPM) positively correlated with the number of spermatozoa injured by snap‐freezing. Eosin‐nigrosin (EN) and propidium iodide readings were correlated with each other, and both provided robust measurements of plasma membrane integrity. A high correlation between expected and measured amounts of acrosome‐intact spermatozoa was found using Fast Green‐Rose Bengal (FG‐RB), Coomassie Blue (CB), and FITC‐PSA stains, and all three methods exhibited comparable results. Likewise, different percentages of UV‐irradiated spermatozoa were accurately assessed for DNA integrity by Toluidine Blue (TB) and sperm chromatin dispersion (SCD) tests. Comparisons between 3,3′‐diaminobenzidine (DAB) and JC‐1 stains also indicated the reliability of the former assay to ascertain gradual increases in spermatozoa with greater mitochondrial function. These data confirmed that different parts of the tamarin spermatozoa can be simply and consistently evaluated by EPM, EN, FG‐RB, CB, TB, and DAB protocols.

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Sperm traits on in vitro production (IVP) of bovine embryos: Too much of anything is good for nothing

Cited by 18 publications

References 52 publications

Reproductive outcomes predicted by phase imaging with computational specificity of spermatozoon ultrastructure

Reproductive outcomes predicted by phase imaging with computational specificity of spermatozoon ultrastructure

Features of the preparation of biological material for genome editing in cattle

Assessment of different sperm functional tests in golden‐headed lion tamarins (Leontopithecus chrysomelas)

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